Bacterias are prokaryotes: they miss a nucleus. Replication,
transcription and translation take place in the same compartment.
In eukaryotes, replication and transcription are separated by
the nuclear wall. In prokaryotes both processes take place in
the same compartment. Bacterial cells show:

In the center of the cytoplasm,
the nucleoid is a region which contains the genomic (here we
should use genophoric instead) DNA (usually one cicular molecule).
This DNA is not associated to histonas nor to any basic bacterial
proteins. However, the eubacterias have a protein of the histona
type, which is called HU in Escherichia coli.The DNA is joined
to the membrana celular during the replicación and the
cell separation. other bacteria, we could found plasmids and
cytoplasmic inclusions.

Mesosom is a small cytoplasmic invagination of the plasma membrane
where ATP is synthetized and which contains the DNA pols for
replication.

Some bacterias show flagels, other, pilis. Pilis are numerous
and short tubes. A Flagel is associated with a basal granule.
A pili can carry the DNA from one cell to the other.

They have ARN and ribosmes for protein synthesis.

The cell wall is rigid and made of molecules typical of bacterias.

--------------------------------------
NB:Escherichia coli (Theodor von Escherich*) is a bacterium
of the intestinal flora. Most species are beneficial. A few pathogenic
species exist nevertheless that liberate enterotoxinas.
E. coli is a bacillus Gram -.
*http://www.whonamedit.com/doctor.cfm/1436.html
http://www.slic2.wsu.edu:82/hurlbert/micro101/pages/
Chap3.html#Bacterial_morphology

Reproduction in bacteria is either sexual or asexual. Sexual
reproduction is an exchange of DNA between two bacterial cells
followed by a modification of the DNA of the recipient bacterium.
For a bacterium there are three ways to exchange DNA:

Transformation: a bacterium takes up DNA directly
from surrounding dead bacteria. DNA enters the cell after a hole
had been made through the cell membrane. A similar process in
eucariotas is called transfection (except in yeast where the
process is also called transformation). Transformation is also
a method used frequently in the laboratory to introduce experimentally
DNA inside bacterias and yeast.

Conjugation and sexduction:
A bacterium contains
a plasmid F (such a bacterium is called F+). This bacterium is
able to build a pilus. The pilus allow the bacterium to communicate
with another bacteria which doesn't possess the plasmid (such
a bacterium is called F-). The exchange of DNA is made through
the pilus. If the plasmid is integrated into the bacterial chromosme
(bacterium HFr), the whole or a part of the chomosome could pass
through the pilus (sexduction).

Transduction: The DNA transfer is made from
one bacteria to the other with the help of a virus (bacteriophage)
infection. This virus behave as a vector. This method is also
used with eucaryotes cells.

In any case,transmission
is llimited by restriction, a mechanism which protects cells
from the introduction of foreign D

It was noticed that when a phage (bacterial virus or bacteriophage)
which parasites one species of E. coli enters in a bacterium
of an other species, it is destroyed: it is said that this phage
is "restricted". The other species produce an endonuclease
which cut the DNA of the phage which was not methylated by a
methylase specific of these other bacteria. The restriction process
is produced by a couple of enzymes: a methylase and an endonuclease.
The methylase and the endonuclease recognize the same site, called
the site of restriction.

type II restriction enzymes
In molecular biology, type II endonucleases are widely used.
The type II methylase puts a methyl on a nucleotido (A or C)
of the site. This ethylation protects this site: indeed, the
endonuclease of the same couple cuts the unmethylated sites.
Any foreign DNA, which is introduced inside the bacterium, is
cut by the endonuclease, because it is not methylated at the
right place. Any DNA produced in this bacterium (cloning) cannot
be cut by the endonuclease of the couple. These two conclusions
are important in molecular biology.

For example, the isoschizomeros Msp I and Hpa II
recognize the same sequence (site of restriction): CCGG.Msp I is sensitive only to the methylation of the carbon
5 of the external cytosine (m5CCGG). Hpa II is sensitive
to the methylation of both cytosines (Cm5CGG, m5CCGG o m5Cm5CGG).

less specific methylases:
Many bacterium exoress less specific methylases like dam and
dcm. You have to know them

dam
dam methylases put a methyl on the nitrogen 6 (m6N) of the molecule
of adenin when located in the sequence GATC. The ADN produced
and extracted from a dam + bacteria cannot be cut by an endonuclease
recognizing a site containing this sequence (Mbo I).

dcm
dcm methylases put a methyl on the carbon 5 (m5) of the molecule
of cytosine located in the sequences CCAGG or CCTGG. The ADN
produced and extracted from a dcm + bacteria cannot be cut by
an endonuclease recognizing a site containing this sequence (Bst
O1).

What to do in practice:
If our recognition site does not contain all or a part the sequences
(GATC) and (CC (A/T) GG), there will be no problem.
But:
The enzymes which recognize these sequences have a variable sensibility
to methylation. Mbo I (sitio: GATC) and Bam H I (sitio: GGATCC)
contain the site dam. Mbo is inhibited and BamH is no.

Restriction and experimental transformation
Transformation requires a bacteria that doesn't destroy, by restriction,
the DNA which is introduced. In eukaryotes, the DNA is often
methylated on CG. When such a DNA is introduced in bacteria expressing
the restriction system Mcr, it is cut into pieces because the
Mcr system cut the DNA which is methylated. Restriction limit
also the transmission of the R plasmid wich communicate the antibiotic
resistence.

The LacZ ([beta]-gal)
product is a polypeptide of 1029 amino acids. It gives rise to
the functional enzyme after tetramerization. Tetramerization
depends on the presence of the 50 first resisdues (N-terminal
region). Deletion of the N-terminal sequence generates a peptide
(omega peptide) that is unable to tetramerize and does not display
enzymatic activity.

The activity of the omega peptide can be fully restored either
in bacteria or in vitro if a small fragment (called alpha peptide)
corresponding to the intact N-terminal portion of the [beta]-gal
is added in trans . The phenomenon is called alpha complementation
and the small N-terminal peptide is called alpha peptide. Special
strains are produced that constitutively express omega peptide.
They don't express enzymatic activity. They can be transformed
by plasmids able to perform alpha complementation. This transformation
restore the missing activity.

Alpha complementation
of LacZ in mammalian cells
Peter Moosmann and Sandro Rusconi
Nucleic Acids Research, Volume 24, Number 6, Pp. 1171-117
Histochemical identification of recombinant clones
The amino-terminal part of the lacZ gene which produces the "alpha"
part of beta-galactosidase has been incorporated into R plasmids.
Bacteria transformed by this plasmid are résistant to
the anitbiotic and perform the alpha complementation. Complementation
is verified by cutivating bacterias with X-gal
(5-bromo-4-chloro-3-indolyl-beta-D- galactoside). When complementation
takes place, colony is blue.

Insertion of cloned DNA into the truncated 5' region of lacZ
gene abolishes this alpha complementation and produce white colony
in the presence of X-gal.

Commercial plasmids have an insertion site (polylinkerintide
the lacZ region.

The ADN of the
procaryotes is organized in groups of genes. Such a group is
called an operon. All the genes of an operon take part in the
same function. In the eucariotas, some protists have operons
(trypanosomes). In this case, the genes of a given operon usually
do not take part in the same function.
Example:
The genes of the operon thr allow the synthesis of the threonine
(thr) when this amino acid is not in the culture medium.

The start site of transcription
(the promoter)
an operator that controls the access of the RNA pol to the promoter
a regulatory gene that codifies a protein that sticks to the
operator. The protein, once tied, the the forward move of the
RNA pol: the transcription is blocked.

The regulatory protein can directly stick to the operator or
only after having fixed the represor.

Then comes the sequences coding the genes of the operon. Every
sequence is separated from the following one by a termination
signal. The transcription will produce a unic mRNA. The translation
will produce each of the codified proteins from this mRNA.

The operons are
inducibles, reprimibles or constitutive. In Escherichia coli
seventy five operons were identified. They control 250 different
structural genes.

Inducible operons are expressed only when the genes that
they contain are necessary. The classic example is the operon
lactose. Lactose is not usually present in the culture medium
of E. coli. The bacterium economizes the expression of the genes
of this operon. But if lactose is present, the operon is induced
and the lactose is used as a metabolite.

Repressible operons are habitually expressed, but they
are repressed when they are not necessary. The operón
trp is not expressed if there is no triptofano in the culture
medium.

In both cases, the operator sequence can fix the regulatory protein.
In the inducibles operons this protein is bound only when the
inductor is absent. In the repressible operons the regulatory
protein is bound when the repressor is present

Constitutive operons are those operons that always express,
and therefore they are regulated neither by inductors nor for
repressors.

The adjustable promoters are promoters acting only when we decide
they should. We insert a gene coding a protein of interest in
a vector. We place an adjustable promoteur in front of this gene.

Indeed, the production of foreign proteins can be toxic for the
bacterium. We begin by allowing the reproduction of bacteria.
Then, when there are enough bacteria, we add an inductor. This
one acts on the promoter: the protein of interest is produced.
When the amount of produced proteins becomes toxic, we stop the
action of the inductor.

The element which controls the operon lac
We make produce by the bacterium or by the vector the protein
"i". We place the promoter and the operator of lac
in front of the inserted gene. The promoter works only when IPTG
is added to the culture medium.

The element which controls the operon trp.
The operon trp allows the synthesis of tryptophane. When there
is tryptophane in the medium this opéron is repressed.
We place the element of controle of trp in front of the gene
to be controlled. The gene is not expressed if there is of the
tryptophane or its equivalent (acid 3-indol-acrilic) in the medium.

The hybrid promoter tac
The promoter tac combines the region-35 of the promoter trp
and-10 of the promoter lac. He is a hybrid promoter. This
promoter is nduced by IPTG and repressed by the glucose.

Promoter thermosensibleIt is possible to use the promoter Pl of the phage lambda
and its mutated repressor CIm. Cim is thermosensible, that is
to say that is destroyed to 42°CAt 30° Clm is stable
and the promoter Pl is inhibited. At 42 °, the inhibitor
is destroyed and the promoter works perfectly well.